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Guaiacol peroxidase zymography for the undergraduate laboratory
Author(s) -
Wilkesman Jeff,
Castro Diana,
Contreras Lellys M.,
Kurz Liliana
Publication year - 2014
Publication title -
biochemistry and molecular biology education
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.34
H-Index - 39
eISSN - 1539-3429
pISSN - 1470-8175
DOI - 10.1002/bmb.20809
Subject(s) - guaiacol , zymography , peroxidase , chromatography , enzyme , gel electrophoresis , chemistry , biochemistry , biological fluids , polyacrylamide gel electrophoresis , enzyme assay , molecular mass , staining , electrophoresis , biology , genetics
This laboratory exercise presents a novel way to introduce undergraduate students to the specific detection of enzymatic activity by electrophoresis. First, students prepare a crude peroxidase extract and then analyze the homogenate via electrophoresis. Zymography, that is, a SDS‐PAGE method to detect enzyme activity, is used to specifically detect peroxidase activity and furthermore, to analyze the total protein profile. After the assay, students may estimate the apparent molecular mass of the enzyme and discuss its structure. After the 4‐h experiment, students gain knowledge concerning biological sample preparation, gel preparation, electrophoresis, and the importance of specific staining procedures for the detection of enzymatic activity. © 2014 by The International Union of Biochemistry and Molecular Biology, 42(5):420–426, 2014.